US5618539A - Stabilized vaccine compositions - Google Patents

Stabilized vaccine compositions Download PDF

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Publication number
US5618539A
US5618539A US08/314,571 US31457194A US5618539A US 5618539 A US5618539 A US 5618539A US 31457194 A US31457194 A US 31457194A US 5618539 A US5618539 A US 5618539A
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United States
Prior art keywords
stabilizer
vaccine composition
lysine
arginine
poliovirus
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Expired - Fee Related
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US08/314,571
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English (en)
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Brent Dorval
Marie Chow
Alexander Klibanov
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Massachusetts Institute of Technology
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Massachusetts Institute of Technology
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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • A61K39/12—Viral antigens
    • A61K39/125—Picornaviridae, e.g. calicivirus
    • A61K39/13—Poliovirus
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • A61K39/12—Viral antigens
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N2770/00—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA ssRNA viruses positive-sense
    • C12N2770/00011—Details
    • C12N2770/32011—Picornaviridae
    • C12N2770/32611—Poliovirus
    • C12N2770/32634—Use of virus or viral component as vaccine, e.g. live-attenuated or inactivated virus, VLP, viral protein
    • Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
    • Y02A—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE
    • Y02A50/00—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE in human health protection, e.g. against extreme weather
    • Y02A50/30—Against vector-borne diseases, e.g. mosquito-borne, fly-borne, tick-borne or waterborne diseases whose impact is exacerbated by climate change

Definitions

  • the trivalent oral polio vaccine (Sabin) is a live-attenuated virus vaccine. It is heat-labile and hence must be stored frozen and used soon after thawing to insure effective immunization against poliomyelitis. Although 1 molar magnesium chloride is an effective stabilizer for the Sabin vaccine, inactivation will still occur if the vaccine thaws during transport or storage. Because of the shortage of adequate refrigeration facilities in underdeveloped and tropical regions, where poliovirus is endemic, the vaccine often cannot be stored frozen and as a consequence the vaccine becomes inactivated. This leads to under-immunization of the populations which are most at risk. Thus, eradication of poliomyelitis depends on the ability to assure cold storage and rapid distribution of poliovirus vaccine. Vaccine formulations with improved stability would circumvent this problem.
  • This invention pertains to stabilized viral vaccines, particularly live viral vaccines for poliomyelitis, comprising an aqueous solution of a live virus and a stabilizing amount of a compound containing at least two amino or imine groups, such as basic amino acids (e.g. lysine).
  • a compound containing at least two amino or imine groups such as basic amino acids (e.g. lysine).
  • These compounds are safe, relatively inexpensive and can be easily added to viral vaccine preparations.
  • the polyamino or imine compound improves the heat stability of the virus in standard tests for viral stability over that of the currently available stabilizer magnesium chloride. This provides more stable live viral vaccine compositions for worldwide distribution and use.
  • FIG. 1 shows stabilization of poliovirus (serotype 1, Mahoney strain) against heat inactivation by 1M amino acids or MgCl 2 .
  • Poliovirus 4 ⁇ 10 8 plaque forming units (PFU, approximately 80 viral particles), was added to 1 ml of 5 mM phosphate buffer, pH 7.0, containing 1M each of L-lysine ( ⁇ ), L-arginine ( ⁇ ), glycine ( ⁇ ), L-alanine (+) or MgCl 2 ( ⁇ ).
  • the resultant solution was placed in 1.4 ml Eppendorf tubes, sealed and submerged in a water bath at 50° C. Aliquots were removed periodically, diluted with with 5 mM phosphate buffer containing 150 mM NaCl, pH 7.0 (PBS), and the titer of infectious poliovirus was followed by plaque assay on HeLa cells.
  • PBS plaque forming units
  • FIG. 2 shows stabilization of poliovirus (serotype 1, Mahoney strain) against heat inactivation by 1 or 2M L-lysine or MgCl 2 .
  • Poliovirus (8 ⁇ 10 8 PFU) was added to 1 ml of 5 mM phosphate buffer, pH 7.5, alone ( ⁇ ) or containing 1M L-lysine ( ⁇ ), 2M L-lysine ( ⁇ ), 1M MgCl 2 ( ⁇ ) or 2M MgCl 2 ( ⁇ ).
  • the resulting solution was placed in 1.4 ml Eppendorf tubes, sealed and submerged in a water bath at 50° C. Aliquots were removed periodically, diluted with PBS, and the titer of infectious poliovirus was followed by plaque assay on HeLa cells.
  • the vaccine compositions of this invention comprise a virus and a compound, containing at least two amino or imine groups, in an amount sufficient to stabilize the virus.
  • the amino or imine compound enhances the stability of the virus against heat inactivation. For example, in standard tests for virus stability at 50° C., the stability of the virus is enhanced at least 10-20 fold by the amino acid lysine.
  • the vaccine compositions are produced by adding the virus and a stabilizing amount of the amino or imine containing compound into a physiologically acceptable aqueous solution.
  • the amino or imine containing compound can be any non-toxic compound containing at least two amino or imine groups.
  • the compounds comprise at least two primary or secondary amino or imine groups separated by a spacer moiety.
  • the size or constituency of the spacer moiety does not appear to be critical.
  • the spacer moiety will consist of a substituted or unsubstituted, linear chain of carbon atoms (heteroatoms such as nitrogen may be included in the chain) ranging from 1 to about 10, preferably from 1 to about 6 atoms.
  • Preferred compounds are the amino acids lysine and arginine or salts (e.g., chloride or acetate) thereof.
  • Some examples of other useful compounds include diaminoethane, 1,3-diaminopropane, 1,4-diaminobutane, and 1,5-diaminopentane.
  • Other stabilizers include compounds which have a nitrogen carrying spacer moiety such as spermidine.
  • polyimines such as poly(ethylenimine) can be used. Mixtures of amino or imine containing compounds can also be used.
  • the stabilizer compound is used in an amount effective to stabilize the virus.
  • concentration of the amino compound is 1-2 molar.
  • the virus can be any virus or mixture of viruses. Generally, the virus will be attenuated.
  • the vaccine compositions can contain any or all of the various types of poliovirus.
  • the preferred vaccines are the trivalent Sabin vaccines which contain types I, II and III of poliovirus.
  • the vaccine compositions will typically be formulated at a pH ranging from about 6 to 8.
  • Magnesium chloride preferably 1 molar, can also be added to the compositions.
  • compositions can combine with the viral components of the compositions.
  • the compositions can contain adjuvants which do not interfere with the activity of the stabilizing compound.
  • Poliovirus serotype 1, Mahoney strain
  • PBS pH 7.2
  • Viral stocks contained approximately 4 ⁇ 10 11 PFU/ml and were stored at 4° C.
  • Approximately 4 ⁇ 10 8 PFU were added to 1 ml of 5 mM phosphate buffer, pH 7.0, alone or containing 1 M L-lysine, D-lysine, L-arginine, glycine, L-alanine, N- ⁇ -acetyl-L-lysine, N- ⁇ -acetyl-L-lysine, L-lysine methyl ester, ethylenediamine, 1,5-diaminopentane, ethylamine, poly(ethylenimine), spermidine or MgCl 2 .
  • the pH of each solution was adjusted to 7.0 with HCl prior to the addition of poliovirus.
  • FIG. 1 demonstrates that lysine and arginine stabilize PV1M 2 to 4 times better than MgCl 2 at all time points, whereas, L-alanine and glycine provide 10 to 10,000 times less stabilization than MgCl 2 during the same period.
  • controls which contained 5 mM phosphate buffer alone at pH 7.0, more than eight orders of magnitude of viral infectivity were lost after 3 hours.
  • FIG. 2 compares stabilization of PVIM by 1 and 2M L-lysine and MgCl 2 at pH 7.0. These data show that L-lysine is 10 and 20 times better than MgCl 2 at stabilizing PVIM after 24 and 48 hours, respectively, at 50° C.

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  • Health & Medical Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Virology (AREA)
  • Pharmacology & Pharmacy (AREA)
  • Animal Behavior & Ethology (AREA)
  • Medicinal Chemistry (AREA)
  • Microbiology (AREA)
  • Mycology (AREA)
  • Chemical & Material Sciences (AREA)
  • Epidemiology (AREA)
  • Immunology (AREA)
  • General Health & Medical Sciences (AREA)
  • Public Health (AREA)
  • Veterinary Medicine (AREA)
  • Medicines Containing Antibodies Or Antigens For Use As Internal Diagnostic Agents (AREA)
  • Medicinal Preparation (AREA)
  • Acyclic And Carbocyclic Compounds In Medicinal Compositions (AREA)
US08/314,571 1989-08-15 1994-09-29 Stabilized vaccine compositions Expired - Fee Related US5618539A (en)

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US08/314,571 US5618539A (en) 1989-08-15 1994-09-29 Stabilized vaccine compositions

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US39399689A 1989-08-15 1989-08-15
US08/314,571 US5618539A (en) 1989-08-15 1994-09-29 Stabilized vaccine compositions

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US39399689A Continuation 1989-08-15 1989-08-15

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US (1) US5618539A (es)
EP (1) EP0487632B1 (es)
JP (1) JP3064413B2 (es)
AT (1) ATE163547T1 (es)
CA (1) CA2065023A1 (es)
DE (1) DE69032102T2 (es)
DK (1) DK0487632T3 (es)
ES (1) ES2113348T3 (es)
WO (1) WO1991002543A1 (es)

Cited By (16)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20060141483A1 (en) * 2004-12-23 2006-06-29 Calton Gary J Stabilization of viral compositions
WO2009111849A1 (en) * 2008-03-10 2009-09-17 Universidade Federal Do Rio De Janeiro - Ufrj Method for stabilized vaccine production
US20090246225A1 (en) * 2003-02-25 2009-10-01 Medimmune, Llc Methods of Producing Influenza Vaccine Compositions
US20100028383A1 (en) * 2005-03-08 2010-02-04 Intervet International B.V. Chemically defined stabiliser
US20110212127A1 (en) * 2008-09-24 2011-09-01 Stabilitech Ltd. Method for Preserving Polypeptides Using a Sugar and Polyethyleneimine
US20110236412A1 (en) * 2008-09-24 2011-09-29 Stabilitech Ltd. Method for Preserving Polypeptides Using a Sugar and Polyethyleneimine
US20120282294A1 (en) * 2003-02-25 2012-11-08 Medimmune, Llc Refrigerator-temperature stable influenza vaccine compositions
US9101607B2 (en) 2010-03-31 2015-08-11 Stabilitech Ltd. Method for preserving alum adjuvants and alum-adjuvanted vaccines
US20150250869A1 (en) * 2012-10-02 2015-09-10 Transgene Sa Virus-containing formulation and use thereof
WO2014204303A3 (en) * 2013-06-17 2015-10-08 De Staat Der Nederlanden, Vert. Door De Minister Van Vws, Ministerie Van Volksgezondheid, Welzijn En Sport Methods for the prevention of aggregation of viral components
US9283270B2 (en) 2012-01-20 2016-03-15 Serum Institute Of India Ltd. Method for stabilization of biological molecules
US10029007B2 (en) 2011-10-05 2018-07-24 Stabilitech Biopharma Ltd Stabilisation of polypeptides
US10206960B2 (en) 2010-03-31 2019-02-19 Stabilitech Biopharma Ltd Stabilisation of viral particles
US10716859B2 (en) 2010-03-31 2020-07-21 Stabilitech Biopharma Ltd Excipients for stabilising viral particles, polypeptides or biological material
US10806783B2 (en) 2014-04-11 2020-10-20 Stabilitech Biopharma Ltd Vaccine compositions
US10980871B2 (en) 2017-05-08 2021-04-20 Iosbio Ltd Vaccine compositions

Families Citing this family (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JP4370451B2 (ja) * 2001-09-28 2009-11-25 大塚製薬株式会社 医薬組成物
WO2003088946A1 (en) * 2002-04-19 2003-10-30 The Regents Of The University Of Michigan Polymer compositions that stabilize and control the release of formaldehyde-treated vaccine antigens
JP6417217B2 (ja) * 2011-06-28 2018-10-31 ロイコケア・アクチェンゲゼルシャフト ウイルスまたは細菌の新規な安定化方法

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US3097143A (en) * 1960-03-30 1963-07-09 Merck & Co Inc Myristyl-gamma-picolinium chloride preserved polio vaccine and its production
US3097142A (en) * 1960-03-30 1963-07-09 Merck & Co Inc Benzyl alcohol stabilized polio vaccines and production thereof
US3128229A (en) * 1961-10-05 1964-04-07 Baylor Medical Foundation Stabilization of viral vaccine and inactivation of adventitious tissue culture contaminants therein
GB1564998A (en) * 1975-03-21 1980-04-16 Connaught Lab Vaccine and stabilizing solution therefor
US4337242A (en) * 1980-02-05 1982-06-29 Merck & Co., Inc. Vaccine stabilizer containing L-glutamic acid and L-arginine
US4338335A (en) * 1980-02-05 1982-07-06 Merck & Co., Inc. Vaccine stabilizer containing L-glutamic acid and L-arginine
EP0065905A1 (fr) * 1981-05-13 1982-12-01 Institut Pasteur Perfectionnements apportés aux agents de stabilisation de virus vivants pour la préparation de vaccins, et vaccins stabilisés contenant lesdits agents de stabilisation

Patent Citations (7)

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US3097143A (en) * 1960-03-30 1963-07-09 Merck & Co Inc Myristyl-gamma-picolinium chloride preserved polio vaccine and its production
US3097142A (en) * 1960-03-30 1963-07-09 Merck & Co Inc Benzyl alcohol stabilized polio vaccines and production thereof
US3128229A (en) * 1961-10-05 1964-04-07 Baylor Medical Foundation Stabilization of viral vaccine and inactivation of adventitious tissue culture contaminants therein
GB1564998A (en) * 1975-03-21 1980-04-16 Connaught Lab Vaccine and stabilizing solution therefor
US4337242A (en) * 1980-02-05 1982-06-29 Merck & Co., Inc. Vaccine stabilizer containing L-glutamic acid and L-arginine
US4338335A (en) * 1980-02-05 1982-07-06 Merck & Co., Inc. Vaccine stabilizer containing L-glutamic acid and L-arginine
EP0065905A1 (fr) * 1981-05-13 1982-12-01 Institut Pasteur Perfectionnements apportés aux agents de stabilisation de virus vivants pour la préparation de vaccins, et vaccins stabilisés contenant lesdits agents de stabilisation

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Davis et al. Microbiology Harper and Row ed. 3rd. Ed. 1980. pp. 1107 1109. *
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Dorval, B.L., et al., "Lysine and Other Diamines Dramatically Stabilize Poliovirus against Thermoinactivation", Biotechnology and Bioengineering 35:1051-1054 (1990).
Dorval, B.L., et al., Lysine and Other Diamines Dramatically Stabilize Poliovirus against Thermoinactivation , Biotechnology and Bioengineering 35:1051 1054 (1990). *
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Cited By (23)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20090246225A1 (en) * 2003-02-25 2009-10-01 Medimmune, Llc Methods of Producing Influenza Vaccine Compositions
US8986705B2 (en) 2003-02-25 2015-03-24 Medimmune, Llc Methods of producing influenza vaccine compositions
US20120282294A1 (en) * 2003-02-25 2012-11-08 Medimmune, Llc Refrigerator-temperature stable influenza vaccine compositions
US8652822B2 (en) * 2003-02-25 2014-02-18 Medimmune, Llc Refrigerator-temperature stable influenza vaccine compositions
US20060141483A1 (en) * 2004-12-23 2006-06-29 Calton Gary J Stabilization of viral compositions
US8778868B2 (en) 2005-03-08 2014-07-15 Intervet International B.V. Chemically defined vaccine stabiliser
US20100028383A1 (en) * 2005-03-08 2010-02-04 Intervet International B.V. Chemically defined stabiliser
TWI398272B (zh) * 2005-03-08 2013-06-11 Intervet Int Bv 化學定義的安定劑
WO2009111849A1 (en) * 2008-03-10 2009-09-17 Universidade Federal Do Rio De Janeiro - Ufrj Method for stabilized vaccine production
US20110236412A1 (en) * 2008-09-24 2011-09-29 Stabilitech Ltd. Method for Preserving Polypeptides Using a Sugar and Polyethyleneimine
US20110212127A1 (en) * 2008-09-24 2011-09-01 Stabilitech Ltd. Method for Preserving Polypeptides Using a Sugar and Polyethyleneimine
US9101607B2 (en) 2010-03-31 2015-08-11 Stabilitech Ltd. Method for preserving alum adjuvants and alum-adjuvanted vaccines
US10716859B2 (en) 2010-03-31 2020-07-21 Stabilitech Biopharma Ltd Excipients for stabilising viral particles, polypeptides or biological material
US10206960B2 (en) 2010-03-31 2019-02-19 Stabilitech Biopharma Ltd Stabilisation of viral particles
US10029007B2 (en) 2011-10-05 2018-07-24 Stabilitech Biopharma Ltd Stabilisation of polypeptides
US9283270B2 (en) 2012-01-20 2016-03-15 Serum Institute Of India Ltd. Method for stabilization of biological molecules
US10111947B2 (en) * 2012-10-02 2018-10-30 Transgene S.A. Virus-containing formulation and use thereof
US20150250869A1 (en) * 2012-10-02 2015-09-10 Transgene Sa Virus-containing formulation and use thereof
KR20160019477A (ko) * 2013-06-17 2016-02-19 드 슈타트 데르 네덜란덴, 베르테겐부어디그트 두어 드 미니스터 반 베이웨이에스, 미니스테리 반 폭스겐트존하이트, 벨지인 엔 스포츠 바이러스 성분의 어그리게이션을 저해하는 방법
US10080793B2 (en) 2013-06-17 2018-09-25 De Staat der Nederlanden, vert, door de minister van VWS, Ministerie van Volksgezonheid, Welzijn en Sport Methods for the prevention of aggregation of viral components
WO2014204303A3 (en) * 2013-06-17 2015-10-08 De Staat Der Nederlanden, Vert. Door De Minister Van Vws, Ministerie Van Volksgezondheid, Welzijn En Sport Methods for the prevention of aggregation of viral components
US10806783B2 (en) 2014-04-11 2020-10-20 Stabilitech Biopharma Ltd Vaccine compositions
US10980871B2 (en) 2017-05-08 2021-04-20 Iosbio Ltd Vaccine compositions

Also Published As

Publication number Publication date
EP0487632A1 (en) 1992-06-03
EP0487632B1 (en) 1998-03-04
ES2113348T3 (es) 1998-05-01
DE69032102D1 (de) 1998-04-09
WO1991002543A1 (en) 1991-03-07
JP3064413B2 (ja) 2000-07-12
JPH04506973A (ja) 1992-12-03
ATE163547T1 (de) 1998-03-15
DE69032102T2 (de) 1998-06-25
DK0487632T3 (da) 1998-05-25
CA2065023A1 (en) 1991-02-16

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